Translation start sequences affect the efficiency of silencing of Agrobacterium tumefaciens T-DNA oncogenes

• Published in: Plant physiology (Bethesda) 2003-11, Vol.133 (3), p.966-977
• Main Authors: Lee, H, Humann, J.L, Pitrak, J.S, Cuperus, J.T, Parks, T.D, Whistler, C.A, Mok, M.C, Ream, L.W
• Format: Article
• Language: English
• Subjects: Agrobacterium tumefaciens; Agrobacterium tumefaciens - genetics; Agrobacterium tumefaciens - growth & development; AMP isopentenyl transferase; Antisense RNA; bacterial diseases of plants; bacterial proteins; bioaccumulation; Codon, Nonsense - genetics; crown galls; disease resistance; DNA; DNA, Bacterial - genetics; Focus Issue: The Agrobacterium-Plant Cell Interaction; gene copy number; gene silencing; Genes; genetic vectors; host plants; host-pathogen relationships; iaaH gene; iaaM gene; indole acetic acid; indole-3-acetamide hydrolase; ipt gene; isopentenyladenine; isopnentenyl-AMP; Messenger RNA; molecular sequence data; Nicotiana tabacum; nucleotide sequences; Oncogenes; Oncogenes - genetics; Phenotype; Plant gall; plant pathogenic bacteria; Plant Tumors - genetics; Plant Tumors - microbiology; Plants; Plants - genetics; Plants - microbiology; Plants, Genetically Modified; posttranscriptional gene silencing; RNA; RNA interference; RNA Interference - physiology; RNA, Antisense - genetics; RNA, Antisense - metabolism; RNA, Messenger - genetics; stop codon; transfer DNA; Transformation, Genetic - genetics; Transgenes; transgenic plants; translation (genetics); translation start sites; tryptophan 5-monoxygenase; Tumors
• ISSN: 0032-0889; 1532-2548
• DOI: 10.1104/pp.103.026534

Agrobacterium tumefaciens oncogenes cause transformed plant cells to overproduce auxin and cytokinin. Two oncogenes encode enzymes that convert tryptophan to indole-3-acetic acid (auxin): iaaM (tryptophan mono-oxygenase) and iaaH (indole-3-acetamide hydrolase). A third oncogene (ipt) encodes AMP isopentenyl transferase, which produces cytokinin (isopentenyl-AMP). Inactivation of ipt and iaaM (or iaaH) abolishes tumorigenesis. Because adequate means do not exist to control crown gall, we created resistant plants by introducing transgenes designed to elicit posttranscriptional gene silencing (PTGS) of iaaM and ipt. Transgenes that elicit silencing trigger sequence-specific destruction of the inducing RNA and messenger RNAs with related sequences. Although PTGS has proven effective against a variety of target genes, we found that a much higher percentage of transgenic lines silenced iaaM than ipt, suggesting that transgene sequences influenced the effectiveness of PTGS. Sequences required for oncogene silencing included a translation start site. A transgene encoding a translatable sense-strand RNA from the 5′ end of iaaM silenced the iaaM oncogene, but deletion of the translation start site abolished the ability of the transgene to silence iaaM. Silencing A. tumefaciens T-DNA oncogenes is a new and effective method to produce plants resistant to crown gall disease.